SUMMARY Damage to myelin from diseases such as multiple sclerosis (MS) results in the disruption of the nerve signal, damage to the axon, and finally degeneration. To date, there are no therapies for repair or remyelination in MS and this fact alone illustrates the greatest hope and unmet need for MS patients. Functional screening for small molecules or biologicals that promote remyelination represents a major hurdle to the identification and development of rational therapeutics for MS. Recently we implemented a novel functional screen using fabricated micropillar arrays to identify anti-muscarinic compounds that greatly enhance oligodendrocyte remyelination (Mei et al., 2014). As many of the promising compounds identified in our initial screen activated or antagonized G-protein coupled receptor (GPCR) targets, in this proposal, we focus screening efforts on GPCR small molecule libraries to identify/confirm/validate receptor targets that either inhibit or promote myelination. We believe that GPCRs represent targetable receptors and pathways for the development of small molecule therapeutics for MS. In this proposal we will: 1. Perform high-throughput screening of GPCR small molecule libraries to identify agonists and antagonists that promote myelination. 2. Identify, confirm and validate novel receptors and pathways responsible for the regulation of oligodendrocyte differentiation and myelination. 3. Investigate the therapeutic implications of activating or blocking specific receptors during development and after demyelination. Overall, we believe that our proposal will not only impart a valuable technical approach but more importantly our data identifies two specific GPCRs, the muscarinic receptor 1 (M1R) that inhibits (Gq) and the kappa opioid receptor (KOR) that promotes (Gi/Go) differentiation and myelination of oligodendrocytes both during development and after demyelination.